Glutathione-based delivery system

ABSTRACT

A delivery system is provided. The delivery system includes a carrier or an active compound and a glutathione or a glutathione derivative grafted thereon. The invention also provides a compound including a moiety comprising a vitamin E derivative or a phospholipid derivative, a polyethylene glycol (PEG) or a polyethylene glycol derivative bonded thereto, and a glutathione (GSH) or a glutathione derivative bonded to the polyethylene glycol or the polyethylene glycol derivative.

This application is a Divisional of U.S. patent application Ser. No.12/244,563, filed Oct. 2, 2008, now U.S. Pat. No. 8,067,380 which is aContinuation-In-Part of U.S. patent application Ser. No. 12/000,261,filed Dec. 11, 2007, now U.S. Pat. No. 7,704,956 which issued Apr. 27,2010. Application Ser. No. 12/000,261 is a Continuation-In-Part of U.S.patent application Ser. No. 11/303,934, filed Dec. 19, 2005, now U.S.Pat. No. 7,446,096, which issued on Nov. 4, 2008. This application alsoclaims priority under 35 U.S.C. §119(a) of Taiwan Patent Application No.94147661, filed on Dec. 30, 2005. The entire contents of each of theabove-identified applications are hereby incorporated by reference.

BACKGROUND OF THE INVENTION

1. Field of the Invention

The invention relates to a biological delivery system, and moreparticularly to a glutathione-based delivery system.

2. Description of the Related Art

The blood brain barrier (BBB) is composed of brain endothelial cellscapable of blocking foreign substances, such as toxin, due to the tightjunction therebetween. Hydrophobic or low-molecular-weight molecules,however, can pass through the BBB via passive diffusion.

Nevertheless, active compounds, such as hydrophilic protein drugs fortreating cerebral or nervous diseases and analgesic peptide drugs actingon the central nervous system, cannot enter brain tissue thereby due totheir large molecular weight or hydrophilicity, resulting indecomposition by enzymes.

Current researches forward various methods of allowing active compoundsto pass through the BBB, including structural modification to increasehydrophobicity of drugs, absorption-mediated transport (AMT) allowingpositive-charged carriers to pass via charge absorption,carrier-mediated transcytosis (CMT) allowing hydrophilic metal ions suchas Na⁺ and K⁺, di-peptides, tri-peptides or glucose to pass viatransporters, and receptor-mediated transcytosis (RMT) allowing macromolecules such as insulin, transferrin, or low-density lipoprotein (LDL)to pass via transcytosis.

Glutathione (GSH) is an endogenous antioxidant. If its concentration inserum is insufficient, some nervous diseases, such as chronic fatiguesyndrome (CFS), may occur.

In 1988, Kiwada Hiroshi provided a liposome capable of accumulation inliver comprising an N-acylglutathione such as N-palmitoylglutathione anda phospholipid such as phosphotidylcholine to target and treat liverdiseases recited in JP63002922.

In 1994, Berislav V. Zlokovic asserted that glutathione (GSH) reachesand passes through the BBB of a guinea pig via a special route, such asGSH-transporter, without decomposition.

In 1995, Berislav V. Zlokovic asserted that glutathione (GSH) exists inbrain astrocyte and endothelial cells with millimolar concentration.

In 1995, Ram Kannan asserted that GSH uptake depends on Na⁺concentration. If Na⁺ concentration is low, GSH uptake from brainendothelial cells may be inhibited. He also pointed Na-dependent GSHtransporter located on the luminal side of the BBB manages GSH uptakeand Na-independent GSH transporter located on the luminal side of theBBB manages efflux of GSH. Additionally, Kannan built a rat hepaticcanalicular GSH transporter (RcGSHT) system using the brains of mice andguinea pigs to analyze cDNA fragments 5, 7, and 11. The results indicatethat fragment 7 represents Na-dependent GSH transporter and fragments 5and 11 represent Na-dependent GSH transporter.

In 1999, Ram Kannan built a mouse brain endothelial cell line (MBEC-4)model simulating BBB situations. The model proved that Na-dependent GSHtransporter is located on the luminal side of the MBEC-4 cell.

In 2000, Ram Kannan asserted that GSH passes through the BBB viaNa-dependent GSH transporter in human cerebrovascular endothelial cells(HCEC) and Na-dependent GSH transporter exists in the luminal plasmamembrane of HCEC.

In 2003, Zhao Zhiyang provided an anti-cancer pro-drug bonded withglutathione s-transferase (GST)/glutathione (GSH) by sulfonamidecovalent bonds to target and treat specific cancer cells after broken ofthe sulfonamide bonds recited in US2003109555. This modification canprotect amino groups of drugs, increase solubility thereof, and alterabsorption and distribution thereof in body.

BRIEF SUMMARY OF THE INVENTION

One embodiment of the invention provides a delivery system comprising acarrier or an active compound, and a glutathione ligand or a glutathionederivative ligand, wherein the glutathione ligand or the glutathionederivative ligand is covalently bound to the carrier or the activecompound, and the glutathione ligand or the glutathione derivativeligand is on an outside surface of the carrier.

One embodiment of the invention provides a compound comprising a moietycomprising a vitamin E, a vitamin E derivative or a phospholipid, apolyethylene glycol or a polyethylene glycol derivative covalently boundthereto, and a glutathione or a glutathione derivative covalently boundto the polyethylene glycol or the polyethylene glycol derivative.

A detailed description is given in the following embodiments withreference to the accompanying drawings.

BRIEF DESCRIPTION OF THE DRAWINGS

The invention can be more fully understood by reading the subsequentdetailed description and examples with references made to theaccompanying drawing, wherein:

FIG. 1 shows a delivery system of the invention.

FIG. 2 shows maximal possible effect (MPE) of various met-enkephalincarriers of the invention.

FIG. 3 shows area under curve (AUC) of various met-enkephalin carriersof the invention.

FIG. 4 shows maximal possible effect (MPE) of various gabapentincarriers of the invention.

FIG. 5 shows area under curve (AUC) of various gabapentin carriers ofthe invention.

FIG. 6 shows serum stability of free met-enkephalin and met-enkephalinin liposomes.

DETAILED DESCRIPTION OF THE INVENTION

The following description is of the best-contemplated mode of carryingout the invention. This description is made for the purpose ofillustrating the general principles of the invention and should not betaken in a limiting sense. The scope of the invention is best determinedby reference to the appended claims.

One embodiment of the invention provides a delivery system comprising acarrier or an active compound, and a glutathione ligand or a glutathionederivative ligand. The glutathione ligand or the glutathione derivativeligand is covalently bound to the carrier or the active compound. Theglutathione ligand or the glutathione derivative ligand is on an outsidesurface of the carrier.

The carrier may comprise a nanoparticle, a polymeric nanoparticle, asolid liquid nanoparticle, a polymeric micelle, a liposome,microemulsion, or a liquid-based nanoparticle. The liposome may compriseat least one of lecithin such as soy lecithin and hydrogenated lecithinsuch as hydrogenated soy lecithin.

The liposome may further comprise cholesterol, water-soluble vitamin E,or octadecyl amine to increase serum resistance or charge amounts. Themolar composition ratio of the liposome may be 0.5-100% of lecithin orhydrogenated lecithin, 0.005-75% of cholesterol or water-soluble vitaminE, and 0.001-25% of octadecyl amine.

The carrier may further encapsulate an active compound with anencapsulation efficiency of about 0.5-100%. The active compound maycomprise small molecule compounds such as gabapentin, peptides such asenkephalin, proteins, DNA plasmids, oligonucleotides, or gene fragmentsand have a molar ratio of about 0.0005-50% in the carrier.

The sulfhydryl group (—SH) of the glutathione ligand may be modified toform the glutathione derivative ligand. The glutathione derivativeligand may have formula (III).

In formula (III), the original sulfhydryl group (—SH) of the glutathioneligand is replaced by —SR. R may comprise C1-10 alkyl or lactoyl(—CO—CH(OH)—CH3).

The glutathione derivative ligand may have formula (IV).

In formula (IV), the original sulfhydryl group (—SH) of the glutathioneligand is replaced by sulfonic acid (—SOOOH).

The carrier or the active compound may target glutathione transportersof organs such as a heart, lung, liver, kidney, or blood brain barrier(BBB).

Specifically, the active compound may pass through the blood brainbarrier, such as brain endothelial cells, with a cell penetration rateof about 0.01-100%.

One embodiment of the invention provides a compound comprising a moietycomprising a vitamin E, a vitamin E derivative or a phospholipid, apolyethylene glycol or a polyethylene glycol derivative covalently boundthereto, and a glutathione or a glutathione derivative covalently boundto the polyethylene glycol or the polyethylene glycol derivative.

The vitamin E derivative may comprise tocopherol derivatives ortocotrienol derivatives and may be α-tocopherol, β-tocopherol,γ-tocopherol, δ-tocopherol, α-tocotrienol, β-tocotrienol, γ-tocotrienol,δ-tocotrienol, α-tocopherol succinate, β-tocopherol succinate,γ-tocopherol succinate, δ-tocopherol succinate, α-tocotrienol succinate,β-tocotrienol succinate, γ-tocotrienol succinate, δ-tocotrienolsuccinate, α-tocopherol acetate, β-tocopherol acetate, γ-tocopherolacetate, δ-tocopherol acetate, α-tocotrienol acetate, β-tocotrienolacetate, γ-tocotrienol acetate, δ-tocotrienol acetate, α-tocopherolnicotinate, β-tocopherol nicotinate, γ-tocopherol nicotinate,δ-tocopherol nicotinate, α-tocotrienol nicotinate, β-tocotrienolnicotinate, γ-tocotrienol nicotinate, δ-tocotrienol nicotinate,α-tocopherol phosphate, β-tocopherol phosphate, γ-tocopherol phosphate,δ-tocopherol phosphate, α-tocotrienol phosphate, β-tocotrienolphosphate, γ-tocotrienol phosphate, or δ-tocotrienol phosphate.

The phospholipid may have formulae (I) or (II).R₁-A₁-  (I)or

In formula (I), A₁ may be sphingosine and R₁ may comprise octanoyl orpalmitoyl. In formula (II), A₂ may be phosphoethanoamine and R₂ maycomprise myristoyl, palmitoyl, stearoyl, or oleoyl.

The polyethylene glycol or the polyethylene glycol derivative may have apolymerization number (n) of about 6-210. The molecular weight of thepolyethylene glycol or the polyethylene glycol derivative may be alteredwith various vitamin E derivatives or phospholipids. For example, whenPEG or its derivative is bound to vitamin E derivatives, it may have amolecular weight of about 300-10,000, when PEG or its derivative isbound to the phospholipid represented by formula (I), it may have amolecular weight of about 750-5,000, and when PEG or its derivative isbound to the phospholipid represented by formula (II), it may have amolecular weight of about 350-5,000.

The polyethylene glycol derivative may comprise carboxylic acid,maleimide, PDP, amide, or biotin.

The sulfhydryl group (—SH) of the glutathione may be modified to formthe glutathione derivative. The glutathione derivative may have formula(III).

In formula (III), the original sulfhydryl group (—SH) of the glutathioneis replaced by —SR. R may comprise C1-10 alkyl or lactoyl(—CO—CH(OH)—CH3).

The glutathione derivative may have formula (IV).

In formula (IV), the original sulfhydryl group (—SH) of the glutathioneis replaced by sulfonic acid (—SOOOH).

Referring to FIG. 1, a delivery system of the invention is illustrated.The delivery system 10 comprises a carrier 20 and a ligand 30 boundthereto. The ligand 30 comprises a moiety 40 comprising a vitamin E, avitamin E derivative or a phospholipid, a polyethylene glycol or apolyethylene glycol derivative 50 bound thereto, and a glutathione or aglutathione derivative 60 bound to the polyethylene glycol and thepolyethylene glycol derivative.

Active compounds, such as proteins, peptides, or small molecules,transported by the targeted carrier with a novel glutathione ligandprovided by the invention can effectively pass through blood brainbarrier by carrier-mediated transcytosis (CMT) or receptor-mediatedtranscytosis (RMT) to treat cerebral or nervous diseases.

EXAMPLE 1

Preparation of TPGS-Glutathione

A stirred solution of N-Cbz Benzyl amino acid (N-Cbz Glutamine, 1.0equiv) and N-hydroxysuccinimide (HOSu, 1.0 equiv) in DME (15 mL) wascooled to 0° C. Dicyclohexylcarbodiimide (DIC, 1.0 equiv) was added andstirred at this temperature for 4 hr. The reaction mixture was allowedto stand for 2 hr in a refrigerator and then filtered.

As expected, the pure compound was obtained in excellent yield (98%)after filtration of the dicyclohexylurea (DCU) formed and evaporation ofthe solvent. The residue was triturated in Et₂O/hexanes, filtered out,and then dried in vacuo to afford a white solid.

The (+)-S-tritylcysteine lithium salt (H-Cys(STrt)-OLi, 1.0 equiv) andsodium carbonate (Na₂CO₃, 5.0 equiv) were dissolved in water (15 mL),and then acetonitrile (CH₃CN) was added followed by the intermediatedproduct obtained in Step-2. The mixture was vigorously stirred at roomtemperature for 3-6 hr until the TLC analysis indicated the absence ofintermediated product in Step-2. The solution was washed with water(2*100 mL) and the organic phase was dried with Na₂SO4, filtered, andconcentrated in vacuo to afford the compound 2.

A stirred solution of compound 2 and N-hydroxysuccinimide (HOSu, 1.0equiv) in DME (15 mL) was cooled to 0° C. Dicyclohexylcarbodiimide (DIC,1.0 equiv) was added and stirred at this temperature for 4 hr. Thereaction mixture was allowed to stand for 2 hr in a refrigerator andthen filtered.

After the DCU and solvent was removed, the glycine lithium salt(H-Gly-OLi, 1.0 equiv) and sodium carbonate (Na₂CO₃, 5.0 equiv) weredissolved in water (15 mL), and then acetonitrile (CH₃CN) was addedfollowed by the intermediated product obtained in Step-4. The mixturewas vigorously stirred at room temperature for 3-6 hr until the TLCanalysis indicated the absence of intermediated product in Step-4. Thesolution was washed with water (2*100 mL) and the organic phase wasdried with Na₂SO₄, filtered, and concentrated in vacuo to afford thecompound 3.

The d-alpha tocopheryl polyethylene glycol 1000 succinate (TPGS-OH) wascoupling with compound 3 via esterification to afford the compound 4.

The compound 4 in methanol (100 mL) was added 10% Pd—C (0.2 times theweight of protected tripeptide-TPGS). The suspension was stirred at roomtemperature for 16 hr under a balloon filled with hydrogen. Thesuspension was filtered through Celite and evaporated, and the residuewas crystallized from ethanol. Then, the compound 5 was obtained.

Triethylsilane (Et₃SiH) and TFA-mediated deprotection of compound 5 inthe presence of CH₂Cl₂ provided the compound 6 (that is GSH-TPGS).

The preparation of TPGS-Glutathione derivatives is similar to theforegoing processes. The distinctions therebetween are simply furthermodifications of the sulfhydryl group (—SH) of the TPGS-Glutathione. Forexample, modifications may be performed, by substitutable groups such asC1-12 alkyl or lactoyl (—CO—CH(OH)—CH3), or oxidization to form sulfonicacid (—SOOOH). The Glutathione and its derivatives are covalently boundto the TPGS with an ester bond.

Preparation of Met-Enkephalin Carrier Solution

0.5 g lipid containing 83.2% soybean phosphatidylcholine (SPC), 4.2%α-tocopherol succinate PEG 1500 (TPGS), 4.2% glutathione-TPGS(GSH-TPGS), and 8.4% cholesterol was placed in a 12.5 mL ZrO₂ mortar.Appropriate amounts of met-enkephalin were dissolved in 10 mM phosphatesolution with pH 7.4 to form a 4% drug solution. 0.5 mL drug solutionand five ZrO₂ beads (10 mm of diameter) were then added to the mortarand ground with 500 rpm for one hour to form a sticky cream. Next, 0.2 gsticky cream and 1.8 mL phosphate solution (10 mM, pH 7.4) were added toa 10 mL flask to hydrate under room temperature for one hour to form acarrier solution containing liposomes encapsulating met-enkephalin. Theconcentration of met-enkephalin in a liposome was 0.56 mg/mL. Theencapsulation efficiency thereof was 33.3%. The mean diameter of thecarrier was 173.1 nm as well as the polydispersity index (PI) was 0.243.

EXAMPLES 2-6

Preparation methods of Examples 2-6 are similar to Example 1. Thedistinctions therebetween are the various carrier compositions. Pleasesee Tables 1 and 2.

TABLE 1 Soy Cho- Octa- Ex- leci- H-soy les- TPGS- decyl Met- amples thinlecithin terol TPGS GSH amine enkephalin 2 10 — 1 — 1 — 0.48 3 10 — 1 —1 1 1.60 4 9 1 1 0.5 0.5 — 1.60 5 9 1 1 0.75 0.25 — 1.60 6 9 1 1 — 1 —1.60

TABLE 2 Mean Met-enkephalin diameter concentration EncapsulationExamples (nm) PI (mg/mL) efficiency (%) 2 162.7 0.227 0.56 31.70 3 161.40.046 4.00 70.33 4 214.1 0.003 3.25 68.85 5 165.3 0.137 3.40 68.48 6214.5 0.116 3.99 80.78

EXAMPLE 7

Preparation of Gabapentin Carrier Solution

0.5 g lipid containing 83.2% soybean phosphatidylcholine (SPC), 4.2%α-tocopherol succinate PEG 1500 (TPGS), 4.2% glutathione-TPGS(GSH-TPGS), and 8.4% cholesterol was placed in a 12.5 mL ZrO₂ mortar.Appropriate amounts of gabapentin were dissolved in 10 mM phosphatesolution with pH 7.4 to form a 10% drug solution. 0.5 mL drug solutionand five ZrO₂ beads (10 mm of diameter) were then added to the mortarand ground with 500 rpm for one hour to form a sticky cream. Next, 0.2 gsticky cream and 1.8 mL phosphate solution (10 mM, pH 7.4) were added toa 10 mL flask to hydrate under room temperature for one hour to form acarrier solution containing liposomes encapsulating gabapentin. Theconcentration of gabapentin in a liposome was 1.08 mg/mL. Theencapsulation efficiency thereof was 35.7%. The mean diameter of thecarrier was 147.7 nm as well as the polydispersity index (PI) was 0.157.

COMPARATIVE EXAMPLE 1

Preparation of Met-Enkephalin Carrier Solution

0.5 g lipid containing 83.2% soybean phosphatidylcholine (SPC), 8.4%α-tocopherol succinate PEG 1500 (TPGS), and 8.4% cholesterol was placedin a 12.5 mL ZrO₂ mortar. Appropriate amounts of met-enkephalin weredissolved in 10 mM phosphate solution with pH 7.4 to form a 4% drugsolution. 0.5 mL drug solution and five ZrO₂ beads (10 mm of diameter)were then added to the mortar and ground with 500 rpm for one hour toform a sticky cream. Next, 0.2 g sticky cream and 1.8 mL phosphatesolution (10 mM, pH 7.4) were added to a 10 mL flask to hydrate underroom temperature for one hour to form a carrier solution containingliposomes encapsulating met-enkephalin. The concentration ofmet-enkephalin in a liposome was 0.57 mg/mL. The encapsulationefficiency thereof was 31.1%. The mean diameter of the carrier was 164.1nm as well as the polydispersity index (PI) was 0.281.

COMPARATIVE EXAMPLES 2-3

Preparation methods of Comparative Examples 2-3 are similar toComparative Example 1. The distinctions therebetween are the variouscarrier compositions. Please see Tables 3 and 4.

TABLE 3 Cho- Comparative Soy H-soy les- Octadecyl Met- Examples lecithinlecithin terol TPGS amine enkephalin 2 10 — 1 1 1 1.60 3 9 1 1 1 — 1.60

TABLE 4 Mean Met-enkephalin Comparative diameter concentrationEncapsulation Examples (nm) PI (mg/ml) efficiency (%) 2 159.7 0.103 3.5870.17 3 149.0 0.168 3.22 69.67

COMPARATIVE EXAMPLE 4

Preparation of Gabapentin Carrier Solution

0.5 g lipid containing 83.2% soybean phosphatidylcholine (SPC), 8.4%α-tocopherol succinate PEG 1500 (TPGS), and 8.4% cholesterol was placedin a 12.5 mL ZrO₂ mortar. Appropriate amounts of gabapentin weredissolved in 10 mM phosphate solution with pH 7.4 to form a 10% drugsolution. 0.5 mL drug solution and five ZrO₂ beads (10 mm of diameter)were then added to the mortar and ground with 500 rpm for one hour toform a sticky cream. Next, 0.2 g sticky cream and 1.8 mL phosphatesolution (10 mM, pH 7.4) were added to a 10 mL flask to hydrate underroom temperature for one hour to form a carrier solution containingliposomes encapsulating gabapentin. The concentration of gabapentin in aliposome was 1.17 mg/mL. The encapsulation efficiency thereof was 38.5%.The mean diameter of the carrier was 155.8 nm as well as thepolydispersity index (PI) was 0.186.

EXAMPLE 8

In Vitro Penetration Rate Test 1 of Met-Enkephalin Liposome

The penetration rate of met-enkephalin was measured using a RBE4/gliomacell model simulating BBB situations. The test results of Examples 1-2(containing glutathione) and Comparative Example 1 (without glutathione)are compared in Table 5.

TABLE 5 Examples Drug dose (μg) Penetration rate (%) SD ComparativeExample 1 182.6 3.4 0.6 Example 1 167.7 9.8 1.3 Example 2 165.2 9.8 1.2

The results indicate that Examples 1 and 2 have an apparently higherpenetration rate (9.8%) of about 2.82 times greater than ComparativeExample 1 (3.4%).

EXAMPLE 9

In Vitro Penetration Rate Test 2 of Met-Enkephalin Liposome

The penetration rate of met-enkephalin was measured using a RBE4/gliomacell model simulating BBB situations. The test results of Example 3(containing glutathione) and Comparative Example 2 (without glutathione)are compared in Table 6.

TABLE 6 Examples Drug dose (μg) Penetration rate (%) SD ComparativeExample 2 250.0 3.55 0.36 Example 3 250.0 6.99 1.43 Example 3 250.0 0.250.03 (glutathione added)

The results indicate that Example 3 has an apparently higher penetrationrate (6.99%) of about 1.96 times greater than Comparative Example 2(3.55%). Additionally, if cells were cultured with glutathione for 30min before Example 3 was performed, the penetration rate thereof waslowered by 0.25% due to the addition of glutathione which occupied theglutathione transporter of the cells to block binding of carriers,reducing drug penetration through the BBB. The result proves that theglutathione carrier provided by the invention passes through the BBB viaglutathione ligand/transporter binding to induce carrier-mediatedtranscytosis (CMT) or receptor-mediated transcytosis (RMT).

EXAMPLE 10

Hot-Plate Test of Met-Enkephalin Liposome

After a laboratory mouse on a 55° C. hot plate was intravenouslyinjected, the analgesic effect on heat-induced pain was evaluated.Referring to FIG. 2, for carriers without glutathione (ComparativeExample 3), 90 min after injection, the maximal possible effect (MPE) ofa 30 mg/mL dose was 13%. For carriers containing glutathione (Example5), 60 min after injection, the maximal possible effect (MPE) of 30mg/mL dose was 37%. Referring to FIG. 3, according to the area undercurve (AUC), Example 5 provides 3.2 times the analgesic effect ofComparative Example 3 and 14.7 times the met-enkephalin solution. Thus,drugs can be safely carried by the carrier with glutathione ligand topass through the BBB to achieve analgesic effect.

EXAMPLE 11

Hot-Plate Test of Gabapentin Liposome

After a laboratory mouse on a 55° C. hot plate was intravenouslyinjected, the analgesic effect on heat-induced pain was evaluated.Referring to FIG. 4, for carriers without glutathione (ComparativeExample 4), 270 min after injection, the maximal possible effect (MPE)of a 10 mg/mL dose was 3.15%. For carriers containing glutathione(Example 7), 180 min after injection, the maximal possible effect (MPE)of a 10 mg/mL dose was 4.47%. Referring to FIG. 5, according to the areaunder curve (AUC), Example 7 provides 1.54 times the analgesic effect ofComparative Example 4 (p<0.005) and 2.76 times the gabapentin solution(p<0.0005). Thus, drugs can be safely carried by the carrier withglutathione ligand to pass through the BBB to achieve analgesic effect.

EXAMPLE 12

Serum Stability Test of Met-Enkephalin Liposome

The carriers provided by Example 5 and fetal bovine serum (FBS) weremixed with 1:1 (v/v) to form a solution. After being placed in a 37° C.water bath for 0, 1, 2, and 4 hours, respectively, the solution wasanalyzed by gel filtration (Sephrox CL-4B, 75 mm×120 mm) and measuredresidual concentration of met-enkephalin in liposomes. The results areshown in FIG. 6.

The results indicate that the concentration of met-enkephalin inliposomes remains 93% above. However, residual concentration of freemet-enkephalin decreases to 2%. It is clear that the carrier provided bythe invention has high serum resistance.

While the invention has been described by way of example and in terms ofpreferred embodiment, it is to be understood that the invention is notlimited thereto. To the contrary, it is intended to cover variousmodifications and similar arrangements (as would be apparent to thoseskilled in the art). Therefore, the scope of the appended claims shouldbe accorded the broadest interpretation so as to encompass all suchmodifications and similar arrangements.

What is claimed is:
 1. A compound, comprising: a first moiety comprisinga vitamin E, a vitamin E derivative or a phospholipid; a second moietycomprising a polyethylene glycol or a polyethylene glycol derivativecovalently bound to the first moiety; and a third moiety comprising aglutathione or a glutathione derivative covalently bound to the secondmoiety.
 2. The compound as claimed in claim 1, wherein the vitamin Ederivative comprises tocopherol derivatives or tocotrienol derivatives.3. The compound as claimed in claim 1, wherein the vitamin E derivativecomprises α-tocopherol, β-tocopherol, γ-tocopherol, δ-tocopherol,α-tocotrienol, β-tocotrienol, γ-tocotrienol, δ-tocotrienol, α-tocopherolsuccinate, β-tocopherol succinate, γ-tocopherol succinate, δ-tocopherolsuccinate, α-tocotrienol succinate, β-tocotrienol succinate,γ-tocotrienol succinate, δ-tocotrienol succinate, α-tocopherol acetate,β-tocopherol acetate, γ-tocopherol acetate, δ-tocopherol acetate,α-tocotrienol acetate, β-tocotrienol acetate, γ-tocotrienol acetate,δ-tocotrienol acetate, α-tocopherol nicotinate, β-tocopherol nicotinate,γ-tocopherol nicotinate, δ-tocopherol nicotinate, α-tocotrienolnicotinate, β-tocotrienol nicotinate, γ-tocotrienol nicotinate,δ-tocotrienol nicotinate, α-tocopherol phosphate, β-tocopherolphosphate, γ-tocopherol phosphate, δ-tocopherol phosphate, α-tocotrienolphosphate, β-tocotrienol phosphate, γ-tocotrienol phosphate, orδ-tocotrienol phosphate.
 4. The compound as claimed in claim 1, whereinthe phospholipid has formula (I):R₁-A₁-  (I) wherein A₁ is sphingosine and R₁ comprises octanoyl orpalmitoyl.
 5. The compound as claimed in claim 1, wherein thephospholipid has formula (II):

wherein A₂ is phosphoethanoamine and R₂ comprises myristoyl, palmitoyl,stearoyl, or oleoyl.
 6. The compound as claimed in claim 1, wherein thepolyethylene glycol or the polyethylene glycol derivative has apolymerization number (n) of about 6-210.
 7. The compound as claimed inclaim 1, wherein when the first moiety is the vitamin E derivative, thepolyethylene glycol or the polyethylene glycol derivative has amolecular weight of about 300-10,000.
 8. The compound as claimed inclaim 4, wherein the polyethylene glycol or the polyethylene glycolderivative has a molecular weight of about 750-5,000.
 9. The compound asclaimed in claim 5, wherein the polyethylene glycol or the polyethyleneglycol derivative has a molecular weight of about 350-5,000.
 10. Thecompound as claimed in claim 1, wherein the polyethylene glycolderivative comprises carboxylic acid, maleimide, PDP, amide, or biotin.